Tumor-associated chromosomal translocations frequently result in the expression of chimeric fusion proteins that behave as aberrant transcription factors. These fusions can act as dominant oncoproteins that are necessary for the formation and maintenance of specific human cancers. EWS/ETS fusions consist of the N-terminus of EWS juxtaposed to the C-terminus of one of five ETS transcription factors, is found in Ewing's family tumors (EFTs). We have found that though both EWS/FLI1 and normal FLU can bind the same genomic loci in vivo, EWS/FLI1 modulates the expression of adjacent genes while normal FLU doesnot. Future rational strategies aimed at inhibiting EWS/ETS fusions depend on elucidating molecular mechanisms through which these proteins modulate target gene expression in comparison to normal ETS counterparts. To address this need the following studies will be performed: (i) relevant cell hosts to study EWS/ETS interactions will be characterized using human EFT derived cell lines and murine embryonal stem (ES) technologies; (ii) EWS/ETS direct target genes whose promoters are bound by three different EWS/ETS proteins will be identified by coupling ChlP/Chip experiments with microarray expression analyses; (iii) transcriptional processes that are modulated by EWS/FLI1 at direct target genes will be investigated. Many human cancers harbor specific mutations in their DMAthat result in the expression of aberrant proteins. In many cases, these proteins play major roles in the genesis and maintenance of these tumors. Antagonizing these abnormal proteins can profoundly inhibit tumor growth. In this proposal we focus on one such abnormal protein, EWS/FLI1, which is found in a subset of highly malignant pediatric cancers. Our goal is that by investigating the molecular mechanisms through which EWS/FLI1 promotes cancer, potent anti- cancer strategies will come to light.